Unlock instant, AI-driven research and patent intelligence for your innovation.

A kind of exendin analog modified by polyethylene glycol and its preparation method and application

A technology of polyethylene glycol and analogs, which is applied in the field of biomedicine and can solve problems such as poor specificity and extended half-life

Active Publication Date: 2019-12-17
ZHEJIANG JIANFENG HANSHENG BIOSCI
View PDF4 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] A technical problem to be solved by the present invention is to overcome the shortcomings of poor specificity when modifying Exendin-4 such as PEGylation, and provide a method for performing site-specific polyethylene glycol (PEG) on Exendin-4 polypeptides and derivatives thereof. The modified product can still maintain the biological activity of Exendin-4 and prolong its half-life

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A kind of exendin analog modified by polyethylene glycol and its preparation method and application
  • A kind of exendin analog modified by polyethylene glycol and its preparation method and application
  • A kind of exendin analog modified by polyethylene glycol and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0099] Example 1. Preparation of linear polyethylene glycol modified Ex4-Cys (C40-PEG-Ex4-Cys)

[0100] A polypeptide (Ex4-Cys) with the following sequence was synthesized using standard polypeptide solid-phase synthesis technology:

[0101] His-Gly-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Leu-Ser-Lys-Gln-Met-Glu-Glu-Glu-Ala-Val-Arg-Leu-Phe-Ile-Glu-Trp- Leu-Lys-Asn-Gly-Gly-Pro-Ser-Ser-Gly-Ala-Pro-Pro-Pro-Pro-Ser-Cys; (Seq ID No.1)

[0102]3, 5, 10, 20 and 50 kDa polyethylene glycols with maleine groups (Nektar, mPEG-pro-pionalaldehyde, mPEG-ALD, 2 kDa, 0.95 mg / ml in 50 mM sodium acetate, pH 5.5) Add respectively 0.5mL Exendin-4 (1 mg / ml in 50mM sodium acetate, pH 5.5), and then add 20mM NaC-NBH3 as reducing agent. The molar ratio of mPEG-ALD to Ex4-Cys is 1:1-2. mPEG-ALD and Ex4-Cys were reacted at 4°C for 2 hours under dark conditions. The reaction was terminated with 0.1% aqueous trifluoroacetic acid (TFA) to obtain PEG-modified Ex4-Cys, named as C40-PEG-Ex4-Cys.

Embodiment 2

[0103] Example 2. Preparation of branched polyethylene glycol modified EX4-Cys (C40-tPEG-Ex4-Cys)

[0104] Add 0.5 mLEx4-Cys (1 mg / ml in 50 mM sodium acetate, pH 5.5), followed by 20 mM NaC-NBH 3 as a reducing agent. The molar ratio of mPEG-ALD to Ex4-Cys is 1:1-2. mPEG-ALD and Ex4-Cys were reacted at 4°C for 2 hours under dark conditions. The reaction was terminated with 0.1% aqueous trifluoroacetic acid (TFA) to obtain branched PEG-modified Ex4-Cys with different molecular weights, named C40-tPEG-Ex4-Cys. Purified C40-tPEG-Ex4-Cys as Figure 1 Shown, its retention time is 18.5 minutes.

Embodiment 3

[0105] Example 3. Analysis of the influence of PEG modification on the physiological activity image of Ex4-Cys

[0106] To detect Exendin-4, PEG-modified Ex4-Cys of different molecular weights reacted with the GLP-1 receptor at a density of 2.5×10 5 The insulin-secreting cells INS-1 were seeded on a 12-well plate, 10 per well 5 cells, and then cultured for 2 days to allow them to stably attach to the bottom of the culture plate. After the cells adhered to the wall, labeled with 125 The buffer of I-Exendin-4 (Exendin-4 derivative extending from amino acid residue 9 and amino acid residue 39) was used to replace the cell culture medium, and a certain amount of buffer was added to form a final concentration of 30 μM. Thereafter, a certain amount of natural Exendin-4 and PEG-modified Ex4-Cys of different molecular weights were added to form a final concentration of 0.001-1000 nM, and incubated at room temperature for 2 hours to enable it to competitively bind to the receptor. W...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
molecular weightaaaaaaaaaa
molecular weightaaaaaaaaaa
molecular weightaaaaaaaaaa
Login to View More

Abstract

The invention relates to a preparation method and application of an exendin analog having undergone site-directed modification by a polyethylene glycol derivative. A product obtained after specific modification of the C terminal of the exendin analog by polyethylene glycol has biological activity similar to the biological activity of exendin-4 and has longer half life in vivo compared with unmodified exendin-4. The polyethylene glycol-modified exendin analog disclosed in the invention is applied to treatment diabetes type II and myocardial infarction and has the advantages of simple preparation, obvious curative effect, long-acting and stable drug effect, easiness in storage, etc. The polyethylene glycol-modified exendin analog is of great significance to promotion of high-efficiency treatment of and development of novel drugs for diabetes and myocardial infarction.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to a polyethylene glycol (PEG) modified Exendin analogue, a preparation method thereof and an application in medicine preparation. Background technique [0002] The function of glucagon peptide-1 (hereinafter referred to as GLP-1) is mainly to induce various biological effects in the body, including stimulating insulin secretion, inhibiting glucagon secretion, promoting satiety, and inhibiting gastrointestinal Peristalsis, increased glucose uptake and weight loss. It has been reported that GLP-1 can effectively prevent pancreatic cell lesions caused by the development of type II diabetes. In non-insulin-dependent diabetes mellitus (NIDDM), GLP-1 can promote the growth of new cells and restore insulin secretion. GLP-1 has the remarkable property of promoting insulin secretion without lowering blood sugar. In addition, the injection of GLP-1 did not cause any toxic side effect...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/575C07K1/107A61K38/22A61K47/60A61P3/10A61P3/04A61P9/10
Inventor 陈小元郎立新牛刚朱雷赵孝斌徐松琳
Owner ZHEJIANG JIANFENG HANSHENG BIOSCI